<p>In large cities, multi-source and multi-plant water supply, while effectively alleviating water shortage, poses notable challenges. In such systems, source switching alters dissolved organic matter (DOM) and microbial communities, whose interaction can destabilize water quality, cause pipe corrosion, form biofilms, and generate harmful by-products, thus threatening supply safety. However, the impact of DOM on microbial communities in water supply systems (WSS) has been less studied. To address this problem, EEM-PARAFAC, bioinformatics, and multivariate statistical analysis were used to get an in-depth understanding of the interaction between DOM and microbial communities. In this study, by analyzing the fluorescence index in the whole process, the DOM from source to tap showed weak humification and a strong autochthonous source. The flocculation process preferentially removed humus components. Moreover, water treatment processes (particularly chlorination) were found to significantly reduce microbial richness by 60.88% and alter community structure composition. Community assembly was largely explained by stochastic processes in subgroups. This stochasticity leads to diverse community structures under different environmental conditions, highlighting the complexity of microbial community formation. In water distribution network, microorganisms were the most stable (lowest AVD index, 0.029), followed by raw water (0.046) and the water treatment plant (0.065). The variations of fulvic-like acid and humic-like acid were considered the key components that need to be paid attention to in the whole process of WSS. This study provided insights into monitoring and managing multi-source multi-water plants WSS schemes.</p>